P737: FUNCTIONAL ROLES OF DDX41 MUTATIONS IN THE DEVELOPMENT OF MYELOID MALIGNANCIES
Authors: A. Kon, M. Nakagawa, K. Kataoka, H. Makishima, M. Nakayama, H. Koseki, Y. Nannya, S. Ogawa
Journal: HemaSphere (2022)

Background:DDX41 is a newly identified leukemia predisposition gene encoding an RNA helicase, whose germline mutations are tightly associated with late-onset myeloid malignancies. Importantly, germline DDX41 mutations were also found in as many as ~3-7 % of sporadic cases with MDS/AML, in which a germline protein-truncating allele is compounded by a somatic missense mutation affecting the helicase domain (p.R525H) in the remaining allele in typical cases. However, little is known about the molecular mechanism by which DDX41 mutations lead to myeloid neoplasms. Aims: This study aimed to clarify the molecular pathogenesis of DDX41 mutated myeloid neoplasms. Methods: We generated mice models carrying conditional/constitutive Ddx41 knock-out (KO) and conditional R525H knock-in (KI) alleles in various combinations. By crossing these mice with Rosa26-CreERT2 transgenic mice, we obtained heterozygous Ddx41 KO (Ddx41+/-), homozygous Ddx41 KO (Ddx41-/-), heterozygous Ddx41 R525H mutation (Ddx41R525H/+), and Ddx41 R525H/KO (Ddx41R525H/-) mice, in which expression of the mutant allele was induced by tamoxifen administration. We then investigated the effect of these alleles on hematopoietic cells. Results: First, we assessed cell intrinsic effects of these Ddx41 alleles, in noncompetitive transplantation experiments. Shortly after tamoxifen administration, most of the recipient mice that were reconstituted with bone marrow (BM) from Ddx41-/- or Ddx41R525H/- mice died within a month after CreERT2 induction due to severe BM failure (BMF). By contrast, the mice transplanted with BM from Ddx41+/- mice showed significantly reduced WBC counts and myeloid skewing in long-term observation. We also assessed the reconstitution capacity of BM cells from different Ddx41 mutant mice using peripheral blood (PB) chimerism after competitive transplantation with Ddx41+/+ cells. The PB donor chimerism of Ddx41-/- and Ddx41R525H/- mice-derived cells was markedly reduced compared to that of Ddx41+/+ cells. In contrast, Ddx41+/- and Ddx41R525H/+ mice-derived cells showed a slightly reduced PB chimerism compared to Ddx41+/+ mice-derived cells. Of interest, when Ddx41R525H/--derived BM cells were co-transplanted with Ddx41+/--derived BM cells, some of the recipient mice showed impaired hematopoiesis, suggesting the presence of a non-cell autonomous effect of Ddx41R525H/- cells on Ddx41+/- cells. Transcriptome analysis of stem cells (Kit+Sca-1-Linlow cells) from different Ddx41 mutant mice revealed significant changes in gene expression and splicing patterns in many genes in all mutant mice, with larger changes for Ddx41R525H/- than Ddx41+/- and Ddx41R525H/+ cells. Notably, Ddx41R525H/--derived stem cells exhibited a significant upregulation of genes involved in innate immunity, including an upregulation of cGAS-STING innate immunity pathways, as well as an enhanced Trp53 pathway, whereas there was a downregulation of genes related to RNA metabolism and ribosome biogenesis. Summary/Conclusion: Our results suggest that monoallelic Ddx41 loss leads to age-dependent impaired hematopoiesis, while compound biallelic loss-of-function and R525 alleles showed early development of severe BM failure, where activated innate immunity and impaired ribosome functions may play important roles. The role of DDX41 mutations in myeloid neoplasms, however, need further evaluation.